ISOLATION AND ANALYSIS OF HUMAN DNA REPAIR GENES

本提案的长期目标是阐明核苷酸 真核系统中的切除修复（NER）机制。 NER是一个 少量的生化途径，保持化学完整性 通过不断的监测和修复过程来修复遗传物质。 NER中的缺陷被发现是导致 几种人类疾病包括着色性干皮病（XP）、科凯恩氏病（Cockayne's 综合征和甲状腺营养不良。此外，NER中的缺陷加速了 致癌作用，并可能参与衰老过程。NER途径 在真核生物中是复杂的， 完成修复过程的基因该项目的重点是 主要是在净入学率的早期阶段， 损伤，为随后的切口准备受损部位， 切口步骤本身。这些早期的步骤似乎涉及大约12至 15个基因这些基因中有相当数量已经被克隆， 这些成就为更详细的生物化学研究奠定了基础。 问题研究拟议项目的具体目标是： XPC基因在NER中的功能2）进一步研究和确定 先前证明的蛋白质-蛋白质的功能相关性 DNA修复因子之间的相互作用，以及3）分析最近分离的 新的基因，以确定它们是否参与NER或 重组修复途径。 这些研究将使用 生物化学和遗传学方法的组合，包括双杂交 系统、无细胞修复测定、检测蛋白质-蛋白质的体外测定 相互作用和定点诱变。

The long term objective of this proposal is the elucidation of nucleotide excision repair (NER) mechanisms in eucaryotic systems. NER is one of a small number of biochemical pathways that maintain the chemical integrity of the genetic material by a constant monitoring and repair process. Defects in NER have been found to be responsible for the development of several human diseases including xeroderma pigmentosum (XP), Cockayne's syndrome, and trichothiodystrophy. In addition, defects in NER accelerate carcinogenesis and may be involved in the aging process. The NER pathway is complex in eucaryotes involving perhaps as many as thirty different genes for the complete repair process. The focus of this project is primarily on the early stages of NER which involve the recognition of damage, priming of the damaged site for subsequent incision and the incision step itself. These early steps appear to involve about twelve to fifteen genes. A significant number of these genes have been cloned and these accomplishments have set the stage for more detailed biochemical studies. The specific aims of the proposed project are to: l) investigate the function of the XPC gene in NER 2) further investigate and determine the functional relevance of previously demonstrated protein-protein interactions among DNA repair factors, and 3) analyze recently isolated novel genes to determine if they have an involvement in either NER or recombinational repair pathways. These studies will be conducted using a combination of biochemical and genetic approaches including the two-hybrid system, cell free repair assays, in vitro assays to detect protein-protein interactions, and site directed mutagenesis.